Our most recent work identified a novel target for the development of anti-HIV therapies. We demonstrated that intracellular host monkey proteins prevent the incorporation of HIV-1 envelope (Env) glycoproteins into nascent virions. We obtained several lines of evidence that suggest that the monkey proteins preclude Env incorporation by interfering with HIV-1 Gag-Env interaction, a precondition for specific Env incorporation into budding virions. Most importantly, these virions, which lack Env, are noninfectious. Thus, Env incorporation represents a novel target for the development of new anti-HIV-1 agents. This observation is reminiscent of the CD4-mediated inhibition of Env incorporation into virions, which is also the subject of drug discovery efforts at Retrovirox. Importantly, we obtained evidence that the Env incorporation defect arises from the inability of Env to reach the cell surface. No drug targeting this critical step of the HIV life cycle is currently available. Moreover, to our knowledge, there has been no attempt to identify small compounds blocking Env cell surface expression and Env incorporation into virions. We recently screened a small number of compounds (1,500) for inhibitors of Env incorporation. This pilot experiment allowed us to identify a few hit compounds, which prevent Env incorporation without apparent toxicity to human cells. These findings demonstrate proof-of-concept that small molecules can block Env incorporation and render virions noninfectious, and make the project proposed here feasible. The objective of this phase I study is complete the screening of a larger library (20,000 compounds) using a specific and sensitive HIV-1 Env incorporation assay. Success in this Phase I application is defined as the milestone identification of at least 10 drug-like compounds with IC50s below 5 5M in HIV replication assays and minimal cytotoxicity (therapeutic index greater than 10). Lead compounds with these profiles will be optimized for potency, specificity and bioavailability in a Phase II project. This second proposal will aim at advancing the preclinical development of these compounds to support human trials. The specific aims of this proposal are: 1) Development of an HIV-1 Envelope incorporation assay, and;2) Screening of small-molecule libraries. PUBLIC HEALTH RELEVANCE: As of the end of 2007, 33 million people worldwide are infected with HIV. Despite advances in antiretroviral therapies, side-effects and the emergence of viruses resistant to currently available drugs pose important problems for the treatment of HIV. We propose to identify and characterize antiretroviral compounds that block HIV through a novel mechanism. These unique compounds could be used for the treatment of HIV infection.